A Host-Directed Approach to the Detection of Infection in Hard-to-Heal Wounds.

Wound infection is traditionally defined primarily by visual clinical signs, and secondarily by microbiological analysis of wound samples. However, these approaches have serious limitations in determining wound infection status, particularly in early phases or complex, chronic, hard-to-heal wounds. Early or predictive patient-derived biomarkers of wound infection would enable more timely and appropriate intervention. The observation that immune activation is one of the earliest responses to pathogen activity suggests that immune markers may indicate wound infection earlier and more reliably than by investigating potential pathogens themselves. One of the earliest immune responses is that of the innate immune cells (neutrophils) that are recruited to sites of infection by signals associated with cell damage. During acute infection, the neutrophils produce oxygen radicals and enzymes that either directly or indirectly destroy invading pathogens. These granular enzymes vary with cell type but include elastase, myeloperoxidase, lysozyme, and cathepsin G. Various clinical studies have demonstrated that collectively, these enzymes, are sensitive and reliable markers of both early-onset phases and established infections. The detection of innate immune cell enzymes in hard-to-heal wounds at point of care offers a new, simple, and effective approach to determining wound infection status and may offer significant advantages over uncertainties associated with clinical judgement, and the questionable value of wound microbiology. Additionally, by facilitating the detection of early wound infection, prompt, local wound hygiene interventions will likely enhance infection resolution and wound healing, reduce the requirement for systemic antibiotic therapy, and support antimicrobial stewardship initiatives in wound care.

Detection of synovial sepsis in horses using enzymes as biomarkers.

BACKGROUND: Synovial sepsis is a commonly occurring, potentially career-ending or even life-threatening orthopaedic emergency. Diagnosis of synovial sepsis is currently primarily based on synovial fluid analysis, which often leaves diagnostic ambiguity due to overlap of clinicopathological parameters between septic and aseptic inflammatory synovitis. OBJECTIVES: To evaluate the reliability of lysozyme (LYS), myeloperoxidase (MPO) and elastase (ELT) as biomarkers for synovial sepsis in horses using a photometric assay to measure increased enzyme activity. STUDY DESIGN: Prospective, single-blinded, analytical, clinical study. METHODS: Equine synovial samples were assigned to one of three groups: (1) healthy controls (n = 10), (2) aseptic (n = 27) and (3) septic synovitis (n = 30). The enzyme activity assays (LYS, MPO and ELT) were compared with standard synovial fluid parameters and broad-range bacterial 16S rDNA PCR. RESULTS: LYS and MPO activities were significantly different between septic synovial samples, and both aseptic and control samples (P < .001, LYS: confidence interval [CI]: 2.25-3.41, resp., 2.21-3.8, MPO: CI 0.752-1.6, resp., 0.639-1.81). LYS achieved a 100% sensitivity and 100% specificity in differentiating between septic and aseptic (cut-off value 751.4) or control (cut-off: 484.6) samples (P < .001). MPO reached 93.33% sensitivity, 100% specificity for distinguishing septic from control (cut-off value: 0.1254) synovial samples and 93.33% sensitivity, 81.48% specificity for discriminating between septic and aseptic (cut-off value: 0.1305) synovial samples (P < .001). ELT activity could not be measured in any synovial sample. Both the LYS and the MPO measurements showed a highly significant correlation with PCR (LYS r = .79, MPO r = .69), synovial leukocyte count (LYS r = .752, MPO r = .571), % neutrophils (LYS r = .751, MPO r = 0.663) and each other (r = .744, all P < .001). MAIN LIMITATIONS: Variation in horses' signalment, affected synovial structures and synovial fluid freezing times may have affected the discriminative power of this study. CONCLUSIONS: Increased MPO and LYS activities allow reliable, rapid diagnosis of synovial sepsis with high sensitivity and specificity.

Feasibility of collecting wound fluid during ongoing negative pressure wound therapy by the use of an additional container - A prospective observational study.

Negative-pressure-wound-therapy is commonly used in clinical routine for wound management. Aim of the present study was to assess the feasibility and safety of using an additional container to collect wound fluid during ongoing negative-pressure-wound-therapy. In this present prospective observational study, patients with negative-pressure-wound-therapy were included. An additional container was inserted in the connecting tube between the wound and the vacuum generating device. The following 3 days, the container was changed daily and replaced by a new one. Further safety outcome parameters were assessed. A questionnaire was answered by the responsible surgeon. Twenty-two patients with negative-pressure-wound-therapy with a median (IQR) age of 58.5 (53.0-70.0) years were included in the present study. In median, the duration of negative-pressure-wound-therapy was 5.0 (4.6-5.5) days. In mean ± SD the collected volume of the wound fluid in millilitres (mL) was on day one 7 ± 4 on day two 8 ± 7 and 10 ± 11 on day three. In one patient, there was <0.1 mL of clear water in the additional container. No safety concerns due to the additional container were observed. This study demonstrates that collecting wound fluid during ongoing negative-pressure-wound-therapy over a time period of 3 days is feasible and safe. No safety concerns were observed.

Chitosan based substrates for wound infection detection based on increased lysozyme activity.

There is a strong need of point-of-care diagnostics for early detection of wound infection. In this study, substrates based on functionalized chitosan were developed for visual detection of elevated lysozyme activity, an infection biomarker in wound fluids. For efficient hydrolysis by lysozyme, N-acetyl chitosan with a final degree of acetylation of around 50% was synthesized. N-acetylated chitosan and a chitosan-starch composite were labeled with structurally different dyes resulting in lysozyme-responsive biomaterials. Incubation with lysozyme in buffer and artificial wound fluid lead to a release of colored hydrolysis products already after 2h incubation. Tests in human wound fluid from infected wounds indicated a clear visual color change after 2.5h compared to control samples. A higher degree of swelling of the chitosan/starch containing substrate led to faster hydrolysis by lysozyme. This study demonstrates the potential of the lysozyme-responsive materials for diagnosis of wound infection and provides different diagnostic substrates for potential incorporation in point-of-care devices.

Bioresponsive polymers for the detection of bacterial contaminations in platelet concentrates.

Bacterial contamination of platelet concentrates (PCs) can lead to fatal transfusion transmitted diseases and is the most abundant infectious risk in transfusion medicine. The storage conditions of PCs provide a good environment for bacterial growth. The detection of these contaminations at an early stage is therefore important to avoid the transfusion of contaminated samples. In this study, bioresponsive polymer (BRP) systems were used for the detection of microorganisms in PCs. The backbone of the polymer consisted of labelled protein (casein), which was demonstrated to be degraded by pure proteases as models and by extracellular enzymes released by contaminating microorganisms. The concomitant colour change was easily visible to the naked eye. To enhance stability, the protein was cross-linked with glycidyl methacrylate (GMA). The cross-linked polymer was easier to handle but was less sensitive than the non-cross-linked material. A contamination of a PC with 10CFU/mL S. aureus was detectable after 24 hours. The visible colour reaction was quantified as a ΔE value according to the CIELab concept. A ΔE value of 21.8 was already reached after 24 hours. Hence, this simple but effective system could prevent transfusion of a contaminated PC.